• Proceedings of the Korean Society of Toxicology Conference
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• 2005.05a
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• pp.85-90
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• 2005

Diesel exhaust Particles (DEPs) are known to induce allergic responses in airway epithelial cells, such as the production of various cytokines via nuclear factor-kappa B ($NF-{\kappa}B$). However. the intracellular signal transduction pathways underlying this phenomenon have not been fully examined. This study showed that DEP induced $NF-{\kappa}B$ activity via transforming growth factor-${\beta}$ activated kinase 1 (TAK1) and $NF-{\kappa}B$-inducing kinase (NIK) in L2 rat lung epithelial cells. DEP induced the $NF-{\kappa}B$ dependent reporter activity approximately two-to three-fold in L2 cells. However, this effect was abolished by the expression of the dominant negative forms of TAK1 or NIK. Furthermore, it was shown that DEP induced TAK1 phosphorylation in the L2 cells. These results suggest that TAK1 and NIK are important mediators of DEP-induced $NF-{\kappa}B$ activation.

• Tuberculosis and Respiratory Diseases
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• v.62 no.4
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• pp.299-307
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• 2007

Background: High mobility group box 1 (HMGB1) is a novel, late mediator of inflammation. This study compared the pro-inflammatory effects of LPS and HMGB1. The transcriptional factors that play an important role in mediating the HMGB1-induced stimulation of IL-8 were also evaluated. Methods: RAW264.7 cells were stimulated with either LPS (100 ng/ml) or HMGB1 (500 ng/ml). The $TNF-{\alpha}$, MIP-2 and $IL-1{\beta}$ levels in the supernatant were evaluated by ELISA at 0, 2, 4, 8, 12 and 24h after stimulation. An acute lung injury was induced by an injection of LPS (5 mg/kg) or HMGB1 (2.5 mg/kg) into the peritoneum of the Balb/c mice. The lung cytokines and MPO activity were measured at 4h (for LPS) or 24h (for HMGB1) after the injection. The transcriptional factor binding sites for NF-IL6, $NF-{\kappa}B$ and AP-1 in the IL-8 promoter region were artificially mutated. Each mutant was ligated with pIL-6luc and transfected into the RAW264.7 cells. One hour after stimulation with HMGB1 (500 ng/ml), the cell lysate was analyzed for the luciferase activity. Results: The expression of MIP-2, which peaked at 8h with LPS stimulation, increased sequentially until 24h after HMGB1 stimulation. An intraperitoneal injection of HMGB1, which induced a minimal increased in $IL-1{\beta}$ expression, provoked the accumulation of neutrophils the lung. A mutation of AP-1 as well as $NF-{\kappa}B$ in the IL-8 promoter region resulted in a lower luciferase activity after HMGB1 stimulation. Conclusion: The proinflammatory effects of HMGB1, particularly on IL-8, are mediated by both $NF-{\kappa}B$ and AP-1.

• The Korean Journal of Physiology and Pharmacology
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• v.13 no.6
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• pp.475-482
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• 2009

Rifampicin is a macrocyclic antibiotic which is used extensively for treatment against Mycobacterium tuberculosis and other mycobacterial infections. Recently, a number of studies have focused on the immune-regulatory effects of rifampicin. Therefore, we hypothesized that rifampicin may influence the TLR2 expression in LPS-activated RAW 264.7 cells. In this study, we determined that rifampicin suppresses LPS-induced TLR2 mRNA expression. The down-regulation of TLR2 expression coincided with decreased production of TNF-$\alpha$ Since NF-${\kappa}B$ is a major transcription factor that regulates genes for TLR2 and TNF-$\alpha$, we examined the effect of rifampicin on the LPS-induced NF-${\kappa}B$ activation. Rifampicin inhibited NF-${\kappa}B$ DNA-binding activity in LPS-activated RAW 264.7 cells, while it did not affect IKK$\alpha/\beta$ activity. However, rifampicin slightly inhibited the nuclear translocation of NF-${\kappa}B$ p65. In addition, rifampicin increased physical interaction between pregnane X receptor, a receptor for rifampicin, and NF-${\kappa}B$ p65, suggesting pregnane X receptor interferes with NF-${\kappa}B$ binding to DNA. Taken together, our results demonstrate that rifampicin inhibits LPS-induced TLR2 expression, at least in part, via the suppression of NF-${\kappa}B$ DNA-binding activity in RAW 264.7 cells. Thus, the present results suggest that the rifampicin-mediated inhibition of TLR2 via the suppression of NF-${\kappa}B$ DNA-binding activity may be a novel mechanism of the immune-suppressive effects of rifampicin.

• Journal of Microbiology and Biotechnology
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• v.18 no.1
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• pp.95-103
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• 2008

Mushrooms are regarded as one of the well-known foods and biopharmaceutical materials with a great deal of interest. Polysaccharide ${\beta}$-glucan is the major component of mushrooms that displays various biological activities such as antidiabetic, anticancer, and antihyperlipidemic effects. In this study, we compared the immunostimulatory potency of polysaccharide fractions, prepared from liquid culture of pine-mushroom Tricholoma matsutake, with a potent immunogen lipopolysaccharide (LPS), and their molecular mechanisms on the functional activation of macrophages. We found that fraction II (TMF-II) was able to comparably upregulate or highly enhance the phenotypic functions of macrophages such NO production and cytokine (IL-$1{\beta}$, IL-6, IL-12, and TNF-${\alpha}$) expression, to LPS. TMF-II triggered the phosphorylation of $I{\kappa}B{\alpha}$, a critical step for NF-${\kappa}B$ activation and translocation. Of the upstream signaling enzymes tested, Src and Akt were thought to be the responsible upstream signaling components in induction of NO production, although TMF-II strongly upregulated the phosphorylation of all MAPK pathways. Therefore, our data suggest that T. matsutake-derived ${\beta}$-glucan may exert its immunostimulating activities with similar potency to LPS via activation of multiple signaling pathways linked to NF-${\kappa}B$ activation.

• Journal of Acupuncture Research
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• v.22 no.3
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• pp.155-167
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• 2005

Objectives : The purpose of this study was to investigate the anti-inflammatory effect of Cobrotoxin on binding affinity of cobrotoxin with P50, $IKK{\alpa}$ and $IKK{\beta}$, activities of NF-${\kappa}B$, Cell viability of astrocyte, expressions of protein molecules of NF-${\kappa}B$ such as P50, P-$1{kappa}B$, $1{\kappa}B$ and iflammation related genes such as Cox-2, iNOS, cPLA2 in the SNP or LPS induced Inflammatory pathway of Rats' astrocytes. Methods : In this study, The expression of cytosolic phospholipase A2, Nitric oxcide, Cyclooxygenase-2 and inducible nitrogen oxide synthase was determined by western blotting with corresponding antibodies, and the generation of NF-${\kappa}B$ was assayed by EMSA method in astrocytes of rats. The Cell viability of astrocytes was determined by MTT assay, and Binding affinity of Cobrotoxin with P50, $IKK{\alpha}$ and $IKK{\beta}$ was assayed by Surface plasmon resonance analysis, and NF-${\kappa}B$ dependent luciferase activity was determined by luciferase analysis, and Uptake of cobrotoxin in astrocytes was identified by Confocal laser scanning microscope Results : 1. Compared with control, LPS-induced NF-${\kappa}B$ DNA binding activity was decreased significantly by 0.1, $0.5{\mu}g/m{\ell}$ of Cobrotoxin in Astrocyte. 2. Compared with control, LPS-induced NF-kB dependent luciferase expression was decreased significantly by 0.1, 0.5 and $1{\mu}g/m{\ell}$ of Cobrotoxin in Astrocyte. 3. Compared with control, SNP induced P50, $I{\kappa}B$ expressions in astrocyte were decreased significantly by 0.1, 0.5 and $1{\mu}g/m{\ell}$ of Cobrotoxin and P-$1{\kappa}B$ expression was decreased significantly by 0.5 and $1{\mu}g/m{\ell}$ of Cobrotoxin. 4. Compared with control, LPS induced P50, $1{\kappa}B$ expressions in astrocyte were decreased significantly by 0.5 and $1{\mu}g/m{\ell}$ of Cobrotoxin. 5. Compared with control, SNP induced Cox-2, iNOS, CPLA2 expressions in astrocyte were decreased significantly by $1{\mu}g/m{\ell}$ of Cobrotoxin. 6. Compared with control, LPS induced Cox-2, cPLA2 expressions in astrocyte were decreased significantly by 0.1, 0.5, $1{\mu}g/m{\ell}$ of Cobrotoxin and iNOS expression was decreased significantly by 0.5, $1{\mu}g/m{\ell}$ of Cobrotoxin. 7. Compared with $0.5{\mu}g/m{\ell}$ of Cobrotoxin, SNP-induced NF-${\kappa}B$ DNA bindins activity in astrocyte was increased significantly by Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 1mM and Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 5mM. 8. Compared with $0.5{\mu}g/m{\ell}$ of Cobrotoxin, LPS-induced NF-${\kappa}B$ DNA binding activity in astrocyte was increased significantly by Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 1mM, Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 5mM, Cobrotoxin $0.5{\mu}g/m{\ell}$with GSH 1mM and Cobrotoxin $0.5{\mu}g/m{\ell}$ with GSH 5mM 9. Compared with $0.1{\mu}g/m{\ell}$ of cobrotoxin, SNP induced P50 expressions in astrocyte were increased significantly by Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 1mM, Cobrotoxin $0.5{\mu}g/m{\ell}$ with DTT 5mM Cobrotoxin $0.5{\mu}g/m{\ell}$ with GSH 1mM and Cobrotoxin $0.5{\mu}g/m{\ell}$ with GSH 5mM. 10. The uptake of the labeled cobrotoxin into the cells was shown under a confocal laser scanning microscope. cobrotoxin was uptaken into the membrane and nucleus of astrocytes. Conclusions : In summary, the present results demonstrate that cobrotoxin directly binds to sulfhydryl group of p50 and IKKS resulting In the reduction of translocation of p50 and IkB release, thereby inhibits activation of NF-${\kappa}B$, and suggest that pico to nanomolar range of cobrotoxin could inhibit the expression of genes in the NF-${\kappa}B$ signal pathway.

• Korean Journal of Food Science and Technology
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• v.51 no.1
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• pp.70-80
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• 2019

The aim of this study was to investigate the effects of red ginseng-derived non-saponin fraction (NSF) on inflammatory responses and monocyte-to-macrophage differentiation in RAW264.7 and THP-1. NSF effectively inhibited inflammatory responses by downregulating nitric oxide (NO) production and protein levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). NSF ($2000{\mu}g/mL$) decreased the levels of NO, iNOS, and COX-2 by 33, 83, and 64%, respectively. NSF inhibited the differentiation of monocyte-to-macrophage by decreasing cell adherence along with downregulation of the cluster of differentiation molecule $11{\beta}$ ($CD11{\beta}$) and CD36. In addition, pro-inflammatory cytokines, such as tumor necrosis factor-alpha, interleukin 6, and monocyte chemoattractant protein 1 (MCP-1), were significantly reduced with NSF treatment. The NSF-mediated inhibition of inflammatory responses was due to the regulation of nuclear factor kappa-light-chain-enhancer of activated B cells ($NF-{\kappa}B$) and nuclear factor (erythroid-derived 2)-like 2 (Nrf2). NSF effectively suppressed the translocation of $NF-{\kappa}B$ into the nucleus, while nuclear Nrf2 and its target protein, heme oxygenase-1, levels were significantly increased.

• Journal of Ginseng Research
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• v.37 no.3
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• pp.315-323
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• 2013

Ginseng is known to have antistress effects. Previously, red ginseng (RG) was shown to repress stress-induced peptidyl arginine deiminase type IV (PADI4) via estrogen receptor ${\beta}$ ($ER{\beta}$) in the brain, thus inhibiting brain cell apoptosis. Moreover, tumor necrosis factor (TNF)-${\alpha}$ plays a critical role in immobilization (IMO) stress. However, the signaling pathway of RG-mediated repressesion of inflammation is not completely understood. In this study, we determined how RG modulated gene expression in stressed brain cells. Since secretion of TNF-${\alpha}$ is modulated via TNF-${\alpha}$ converting enzyme (TACE) and nuclear factor (NF)-${\kappa}B$, we examined the inflammatory pathway in stressed brain cells. Immunohistochemistry revealed that TACE was induced by IMO stress, but RG repressed TACE induction. Moreover, PADI4 siRNA repressed TACE expression compared to the mock transfected control suggesting that PADI4 was required for TACE expression. A reporter assay also revealed that $H_2O_2$ oxidative stress induced NF-${\kappa}B$ in neuroblastoma SK-N-SH cells, however, RG pretreatment repressed NF-${\kappa}B$ induction. These findings were supported by significant induction of nitric oxide and reactive oxygen species (ROS) by oxidative stress, which could be repressed by RG administration. Taken together, RG appeared to repress stress-induced PADI4 via TACE and NF-${\kappa}B$ in brain cells thus preventing production of ROS and subsequently protecting brain cells from apoptosis.

• Natural Product Sciences
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• v.16 no.4
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• pp.265-270
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• 2010

The methanol extract of Zanthoxylum rhetsa (MZRR) were evaluated for its ability to suppress the formation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. MZRR presented an inhibition of LPS-induced production of nitric oxide (NO) and prostaglandin $E_2$ ($PGE_2$) in RAW 264.7 macrophages. Western blotting and RT-PCR analyses demonstrated that MZRR significantly inhibited the protein and mRNA expressions of iNOS and COX-2 in LPS-activated macrophages in a dose-dependent manner. LPS-induced COX-2, iNOS, and nuclear factor kappa beta (NF-${\kappa}B$) activity were also decreased in the presence of MZRR. The production of tumor necrosis factor-$\alpha$ (TNF-$\alpha$), the mRNA expression levels of pro-inflammatory cytokines, including TNF-$\alpha$ and IL-$1{\beta}$, were reduced after MZRR administration in a dose dependent-manner. These results suggest that the MZRR extract involved in the inhibition of iNOS and COX-2 via the NF-${\kappa}B$ pathway, revealing a partial molecular basis for anti-inflammatory properties of the MZRR extract.

• Nutrition Research and Practice
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• v.11 no.4
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• pp.265-274
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• 2017

BACKGROUND/OBJECTIVES: Nelumbo leaves have been used in traditional medicine to treat bleeding, gastritis, hemorrhoids, and halitosis. However, their mechanisms have not been elucidated. MATERIALS/METHODS: The present study prepared two Nelumbo leaf extracts (NLEs) using water or 50% ethanol. Inflammatory response was induced with LPS treatment, and expression of pro-inflammatory mediators (inducible nitric oxide synthase, cyclooxygenase-2, tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$), interleukin (IL)-$1{\beta}$, and IL-6 and nitric oxide (NO) and prostaglandin $E_2$ ($PGE_2$) productions were assessed. To determine the anti-inflammatory mechanism of NLEs, we measured nuclear factor-${\kappa}B$ (NF-${\kappa}B$) activity. Major metabolites of NLEs were also analyzed and quantified. RESULTS: NLEs effectively reduced the expression and productions of pro-inflammatory mediators such as IL-$1{\beta}$, IL-6, TNF-${\alpha}$, $PGE_2$, and NO. NLEs also reduced NF-${\kappa}B$ activity by inhibiting inhibitor of NF-${\kappa}B$ phosphorylation. Both extracts contained catechin and quercetin, bioactive compounds of NLEs. CONCLUSIONS: In this study, we showed that NLEs could be used to inhibit NF-${\kappa}B$-mediated inflammatory responses. In addition, our data support the idea that NLEs can ameliorate disease conditions involving chronic inflammation.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2019.04a
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• pp.94-94
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• 2019

In this study, we elucidated the anti-inflammatory mechanisms of leaves extracts from Rodgersia podophylla (RPL) in RAW264.7 cells. RP-L significantly inhibited the production of the proinflammatory mediators such as NO, iNOS, IL-$1{\beta}$ and IL-6 in LPS-stimulated RAW264.7 cells. RPL increased HO-1 expression in RAW264.7 cells, and the inhibition of HO-1 by ZnPP reduced the inhibitory effect of RPL against LPS-induced NO production in RAW264.7 cells. Inhibition of p38, ROS and $GSK3{\beta}$ attenuated RPL-mediated HO-1 expression. Inhibition of ROS inhibited p38 phosphorylation and $GSK3{\beta}$ expression induced by RPL. In addition, inhibition of $GSK3{\beta}$ blocked RPL-mediated p38 phosphorylation. RPL induced nuclear accumulation of Nrf2, and Inhibition of p38, ROS and $GSK3{\beta}$ abolished RPL-mediated nuclear accumulation of Nrf2. Furthermore, RPL blocked LPS-induced degradation of $I{\kappa}B-{\alpha}$ and nuclear accumulation of p65. RP-L also attenuated LPS-induced phosphorylation of ERK1/2 and p38. Our results suggest that RPL exerts potential antiinflammatory activity by activating ROS/$GSK3{\beta}$/p38/Nrf2/HO-1 signaling and inhibiting NF-${\kappa}B$ and MAPK signaling in RAW264.7 cells. These findings suggest that RPL may have great potential for the development of anti-inflammatory drug.